U.S. patent application number 10/012729 was filed with the patent office on 2003-06-12 for multi-channel reagent dispensing apparatus and method.
This patent application is currently assigned to Biosearch Technologies, Inc.. Invention is credited to Baltzley, Arnie, Cook, Ronald M., Dill, Rand.
Application Number | 20030109060 10/012729 |
Document ID | / |
Family ID | 21756416 |
Filed Date | 2003-06-12 |
United States Patent
Application |
20030109060 |
Kind Code |
A1 |
Cook, Ronald M. ; et
al. |
June 12, 2003 |
Multi-channel reagent dispensing apparatus and method
Abstract
Embodiments of the present invention are directed to an improved
chemical synthesis apparatus for performing chemical synthesis such
as nuclei acid synthesis in a plurality of reaction wells or cells
in an efficient manner. The apparatus employs dispenser heads that
each include a cluster of nozzles which are fluidicly coupled to a
plurality of reagent sources for dispensing different reagents
through the single dispenser head. Because each dispenser head is
capable of dispensing a plurality of different reagents, the
apparatus can perform simultaneous synthesis in a plurality of
cells at a high throughput without complex and elaborate control of
movement of the dispenser heads relative to the cells.
Inventors: |
Cook, Ronald M.; (Novato,
CA) ; Dill, Rand; (Corte Madera, CA) ;
Baltzley, Arnie; (San Anselmo, CA) |
Correspondence
Address: |
Townsend and Townsend and Crew LLP
Two Embarcadero Center, 8th Floor
San Francisco
CA
94111
US
|
Assignee: |
Biosearch Technologies,
Inc.
81 Digital Drive
Novato
CA
94949
|
Family ID: |
21756416 |
Appl. No.: |
10/012729 |
Filed: |
December 7, 2001 |
Current U.S.
Class: |
436/180 ;
422/501 |
Current CPC
Class: |
B01J 2219/00389
20130101; B01J 2219/00689 20130101; Y10T 436/119163 20150115; Y10T
436/2575 20150115; B01J 2219/00315 20130101; B01J 2219/00423
20130101; G01N 35/1065 20130101; C40B 60/14 20130101; B01J
2219/0036 20130101; G01N 35/1002 20130101; B01J 2219/00596
20130101; B01J 2219/00414 20130101; B01L 3/0203 20130101; B01J
19/0046 20130101; B01J 2219/00317 20130101; B01J 2219/00585
20130101; C40B 40/06 20130101; B01J 2219/0059 20130101; B01J
2219/00722 20130101; Y10T 436/114165 20150115; Y10T 436/11
20150115 |
Class at
Publication: |
436/180 ;
422/100 |
International
Class: |
B01L 003/02 |
Claims
What is claimed is:
1. A multi-channel reagent dispenser head for introducing a
plurality of reagents into a reaction site, the dispenser head
comprising: a dispenser head body having a dispensing end which is
configured to dispense a plurality of reagents from a plurality of
reagent sources; and a group of nozzles including a plurality of
reagent dispensing nozzles which are fluidicly coupled with the
plurality of reagent sources, the group of nozzles being clustered
to provide a plurality of nozzle outlets in the dispenser head body
to introduce reagents from the plurality of reagent sources through
the dispensing end of the dispenser head body into the reaction
site.
2. The dispenser head of claim 1 wherein the plurality of reagent
dispensing nozzles are each separately coupled with one of the
plurality of reagent sources.
3. The dispenser head of claim 2 wherein the plurality of reagent
dispensing nozzles are separately coupled with reagent sources of
bases A, C, G, T, and an activator.
4. The dispenser head of claim 2 wherein the plurality of reagent
dispensing nozzles are separately coupled with reagent sources of
acid deblock, oxidizers, and capping agents.
5. The dispenser head of claim 1 wherein the group of nozzles
include a wash nozzle which is fluidicly coupled with a source of
wash solvent.
6. The dispenser head of claim 5 wherein the wash solvent comprises
acetonitrile.
7. The dispenser head of claim 1 wherein the group of nozzles
include a vacuum nozzle which is coupled to a vacuum source.
8. The dispenser head of claim 7 wherein the nozzle outlet of the
vacuum nozzle is disposed proximal of the nozzle outlets of the
reagent dispensing nozzles.
9. The dispenser head of claim 1 wherein the dispensing end of the
dispenser head body has a maximum dimension of about 9 mm.
10. The dispenser head of claim 1 wherein the nozzles each have an
outer diameter of less than about {fraction (1/16)} inch.
11. A multi-channel reagent dispenser apparatus for introducing a
plurality of reagents into a plurality of reaction sites, the
apparatus comprising: a plurality of reagent sources; a plurality
of reagent dispensing nozzles each coupled with one of the
plurality of reagent sources; a plurality of dispenser heads, each
dispenser head including a dispenser head body having a dispensing
end which is configured to dispense a plurality of reagents from
the plurality of reagent sources, each dispenser head body having
therein a group of nozzles being clustered to provide a plurality
of nozzle outlets in the dispenser head body to introduce reagents
from the plurality of reagent sources through the dispensing end of
the dispenser head body into one of the reaction sites, the group
of nozzles including more than one reagent dispensing nozzle from
the plurality of reagent dispensing nozzles.
12. The apparatus of claim 11 further comprising a plurality of
reagent dispensing nozzle valves each coupled with one of the
reagent dispensing nozzles to control reagent flow from the reagent
sources to the reagent dispensing nozzles.
13. The apparatus of claim 11 further comprising at least one
source of wash solvent and at least one wash nozzle coupled with
the at least one source of wash solvent, wherein the group of
nozzles clustered in each dispenser head body include at least one
wash nozzle.
14. The apparatus of claim 13 further comprising at least one wash
nozzle valve each coupled with one of the at least one wash nozzle
to control wash solvent flow from the at least one source of wash
solvent to the at least one wash nozzle.
15. The apparatus of claim 11 wherein the group of nozzles
clustered in each dispenser head body include a vacuum nozzle which
is coupled to a vacuum source.
16. The apparatus of claim 15 further comprising a vacuum nozzle
valve coupled with the vacuum nozzle to control vacuum flow through
the vacuum nozzle.
17. The apparatus of claim 11 wherein the plurality of dispenser
heads comprise at least one first dispenser head and at least one
second dispenser head, each first dispenser head having therein a
cluster of first nozzles being coupled with a first set of the
plurality of reagent sources, each second dispenser head having
therein a cluster of second nozzles being coupled with a second set
of the plurality of reagent sources which are different from the
first set of reagent sources.
18. The apparatus of claim 17 wherein the first set of reagent
sources comprise bases A, C, G, T, and an activator.
19. The apparatus of claim 17 wherein the second set of reagent
sources comprise acid deblock, oxidizers, and capping agents.
20. The apparatus of claim 17 further comprising a first actuator
configured to move each of the at least one first dispenser head
from one reaction site to another reaction site to introduce
reagents from the first set of reagent sources into the reaction
sites.
21. The apparatus of claim 20 further comprising a second actuator
configured to move each of the at least one second dispenser head
from one reaction site to another reaction site to introduce
reagents from the second set of reagent sources into the reaction
sites.
22. The apparatus of claim 21 further comprising a controller
coupled with the first and second actuators to automatically
control movements of the at least one first dispenser head and the
at least one second dispenser head to introduce reagents from the
first and second sets of reagent sources separately into the
reaction sites.
23. The apparatus of claim 22 further comprising a plurality of
reagent dispensing nozzle valves each coupled with one of the
reagent dispensing nozzles, wherein the controller is coupled with
the reagent dispensing nozzle valves to control reagent flow from
the reagent sources to the first nozzles in the at least one first
dispenser head and to the second nozzles in the at least one second
dispenser head.
24. The apparatus of claim 17 wherein the dispensing end of each
dispenser head body has a maximum dimension of about 9 mm.
25. The apparatus of claim 24 wherein the plurality of dispenser
heads comprise a plurality of first dispenser heads and a plurality
of second dispenser heads, wherein the first dispenser heads are
spaced about 9 mm apart, and wherein the second dispenser heads are
spaced about 9 mm apart.
26. The apparatus of claim 11 wherein the plurality of reagent
sources each are delivered to the reagent dispensing nozzles by
pressurization with an inert gas.
27. The apparatus of claim 11 wherein the reaction sites are
evacuated under vacuum assist.
28. The apparatus of claim 11 wherein the reaction sites are
disposed in an array provided in a plurality of vacuum trays which
are formed on a single block.
29. The apparatus of claim 11 wherein the apparatus is disposed in
an inert environment.
30. A method for introducing a plurality of reagents into a
plurality of reaction sites, the method comprising: providing a
plurality of reagent sources; providing a plurality of reagent
dispensing nozzles each coupled with one of the plurality of
reagent sources; providing a plurality of dispenser heads, each
dispenser head including a dispenser head body having a dispensing
end, each dispenser head body having therein a group of nozzles
being clustered to provide a plurality of nozzle outlets in the
dispenser head body, the group of nozzles including more than one
reagent dispensing nozzle from the plurality of reagent dispensing
nozzles; and controlling flows of reagents from the plurality of
reagent sources through the plurality of reagent dispensing nozzles
to dispense a plurality of reagents via the group of nozzles
clustered in each dispenser head body through the dispensing end of
the dispenser head body into one of the reaction sites.
31. The method of claim 30 wherein the flows of reagents through
each dispenser head body are controlled by operating a plurality of
reagent dispensing valves each coupled with one of the reagent
dispensing nozzles based on flow rates to dispense preset amounts
of reagents via the group of nozzles clustered in each dispenser
head body at preset times through the dispensing end of the
dispenser head body into one of the reaction sites.
32. The method of claim 30 wherein the flows of reagents through
each dispenser head body are controlled to provide one reagent at a
time through the dispenser head body.
33. The method of claim 30 further comprising: providing at least
one source of wash solvent and at least one wash nozzle coupled
with the at least one source of wash solvent, wherein the group of
nozzles clustered in each dispenser head body include at least one
wash nozzle; and dispensing wash solvent through the at least one
wash nozzle in each dispenser head body at preset times.
34. The method of claim 30 further comprising: providing a vacuum
nozzle in the group of nozzles clustered in each dispenser head
body; and drawing a vacuum through the vacuum nozzle in each
dispenser head body between dispensing different reagents through
the dispenser head body.
35. The method of claim 30 wherein the plurality of dispenser heads
comprise at least one first dispenser head and at least one second
dispenser head, each first dispenser head having therein a cluster
of first nozzles being coupled with a first set of the plurality of
reagent sources, each second dispenser head having therein a
cluster of second nozzles being coupled with a second set of the
plurality of reagent sources which are different from the first set
of reagent sources.
36. The method of claim 35 further comprising moving each of the at
least one first dispenser head from one reaction site to another
reaction site to introduce reagents from the first set of reagent
sources into the reaction sites.
37. The method of claim 36 further comprising moving each of the at
least one second dispenser head from one reaction site to another
reaction site to introduce reagents from the second set of reagent
sources into the reaction sites.
38. The method of claim 37 wherein the at least one first dispenser
head and the at least one second dispenser head are moved
automatically by computer control.
39. The method of claim 37 wherein the plurality of dispenser heads
comprise a plurality of first dispenser heads, and further
comprising controlling flows through the first nozzles in the first
dispenser heads to dispense reagents from the first set of reagent
sources to separate reaction sites simultaneously.
40. The method of claim 37 wherein the plurality of dispenser heads
comprise a plurality of second dispenser heads, and further
comprising controlling flows through the second nozzles in the
second dispenser heads to dispense reagents from the second set of
reagent sources to separate reaction sites simultaneously.
Description
BACKGROUND OF THE INVENTION
[0001] The present invention relates generally to solid phase
processing and, more particularly, to an apparatus for dispensing
reagents and other fluids to a plurality of reaction sites for
solid phase processing including solid phase synthesis of complex
chemicals such as oligonucleotides and the like.
[0002] A variety of separative, synthetic, and enzymatic or
otherwise catalytic processes use beds of particulate material with
transport of reactants, reagents and products or eluants in
solution through the bed. In addition, many reactions are known in
which the products are separated by concentration in one of two or
more phases. These processes include, among others, ion exchange
chromatography, gel filtration, ion exclusion chromatography,
affinity chromatography, separations based on hydrophobicity,
purification based on hybridization, peptide synthesis,
oligonucleotide synthesis, and polysaccharide synthesis including
combinations of the last three. These processes may be carried out
on a small scale for analytical purposes or process design, and are
then often scaled up for preparative work. In nearly all examples
the solid phase particulates are packed in a closed column with a
porous frit on the lower end, an optional frit at the top, and with
fluid-connections at both ends so that liquid can flow in either
direction through the bed. To achieve efficiency and high
resolution with solid phase supports, all volume elements of all
fluids should flow through paths of identical composition and
nearly identical length, and all particles in the bed should be
exposed to the same succession of liquids under the same
conditions.
[0003] In solid phase systems, some interaction occurs between the
solutes running through the bed and the particles composing the
bed. This interaction may be based on secondary forces (ionic,
hydrophobic, or on immunochemical interactions, or base pairing) or
primary valencies as when amino acids or nucleotides are added to a
growing chain on the solid phase support, or when immobilized
enzymes cleave substrates flowing through the bed, or when enzymes
in solution react with substrates attached to the packing. In
addition, solvents or reagents of successively differing
composition which dissociate adsorbed or otherwise attached bound
molecular species, or which cleave off protective groups, or
compounds including polymers which have been synthesized on the
support may be made to flow through the support. The dissociated or
cleaved substances then are free to flow out of the bed in flowing
liquid.
[0004] In particular, nucleic acid synthesis (generally referred to
as "DNA synthesis") is the process of constructing synthetic
single-stranded oligonucleotide through linking of nucleotide, the
basic building blocks for DNA. In an automated system, the various
steps are carried out by a reagent delivery system which dispenses
a number of chemical reagents in a predetermined sequence in a
cycle into a synthesis reaction column containing the solid-phase
support, according to instructions from the system controller or
computer. After the desired number of cycles have been completed,
the synthesized oligonucleotide is separated from the reaction
column and collected in a vial. This step is generally referred to
as "cleavage". The oligonucleotide may further be subject to a step
generally referred to as "deprotection" to complete isolation of
the oligonucleotide. In a process for synthesizing polynucleotides
on a solid support, the solid support traditionally consists of
glass beads of controlled porosity (CPG) or, more generally, of
particles of a functionalized inorganic or organic polymer.
[0005] The isolation of oligonucleotide involves the treatment of
the solid bound oligonucleotide with a cleavage and/or deprotection
reagent. Typically, this reagent is concentrated ammonia solution
in water but can be other homogeneous or heterogeneous solutions of
appropriate bases, alcohols and water. The cleavage and
deprotection process is typically performed in two steps. The
cleavage of the oligonucleotide is performed at room temperature
for approximately one hour before decanting the mixture into a
pressure-sealable vessel for extended higher temperature treatment
to effect the removal of secondary protecting groups on the
synthetic oligonucleotide. This two step process reduces the
quantity of support related contaminants in the final isolated
product.
[0006] The use of a single nozzle for delivering different reagents
into a reaction site, well, or column is not feasible because the
nozzle will need to be cleaned or flushed out between reagents to
avoid contamination, resulting in a high cost and a low throughput.
In one conventional chemical synthesis system, a plurality of
reagent dispensing nozzles are arranged in a linear array, and the
plate containing the reaction cell(s) or column(s) is moved under
the linear array to receive reagents from the dispensing nozzles
one at a time. The throughput remains low.
[0007] Another synthesis apparatus is disclosed in U.S. Pat. Nos.
5,814,700, 5,837,858, and 6,001,311 employing an array of nozzles.
A transport mechanism aligns the reaction wells and selected
nozzles for deposition of the liquid reagent into the selected
reaction wells. Elaborate manipulation of the transport mechanism
is used to dispense reagents from the various nozzles into the
various reaction wells in sequence to provide simultaneous
synthesis in the reaction wells. The throughput is still relatively
low because each nozzle can dispense only one reagent.
BRIEF SUMMARY OF THE INVENTION
[0008] Embodiments of the present invention are directed to an
improved chemical synthesis apparatus for performing chemical
synthesis such as nuclei acid synthesis in a plurality of reaction
wells or cells in an efficient manner. The apparatus employs
dispenser heads that each include a cluster of nozzles which are
fluidicly coupled to a plurality of reagent sources for dispensing
different reagents through the single dispenser head. Because each
dispenser head is capable of dispensing a plurality of different
reagents, the apparatus can perform simultaneous synthesis in a
plurality of cells at a high throughput without complex and
elaborate control of movement of the dispenser heads relative to
the cells.
[0009] In accordance with an aspect of the present invention, a
multi-channel reagent dispenser head for introducing a plurality of
reagents into a reaction site comprises a dispenser head body
having a dispensing end which is configured to dispense a plurality
of reagents from a plurality of reagent sources. A group of nozzles
include a plurality of reagent dispensing nozzles which are
fluidicly coupled with the plurality of reagent sources. The group
of nozzles are clustered to provide a plurality of nozzle outlets
in the dispenser head body to introduce reagents from the plurality
of reagent sources through the dispensing end of the dispenser head
body into the reaction site.
[0010] In some embodiments, the plurality of reagent dispensing
nozzles are each separately coupled with one of the plurality of
reagent sources. The plurality of reagent dispensing nozzles may be
separately coupled with reagent sources of building block elements
such as bases A, C, G, T, and an activator such as tetrazole.
Alternatively, the plurality of reagent dispensing nozzles may be
separately coupled with reagent sources of acid deblock, oxidizers,
and capping agents. The group of nozzles desirably include a wash
nozzle which is fluidicly coupled with a source of wash solvent.
The wash solvent may comprise acetonitrile. The group of nozzles
desirably include a vacuum nozzle which is coupled to a vacuum
source. The nozzle outlet of the vacuum nozzle may be disposed
proximal of the nozzle outlets of the reagent dispensing nozzles.
In specific embodiments, the dispensing end of the dispenser head
body has a maximum dimension of about 9 mm. The nozzles each have
an outer diameter of less than about {fraction (1/16)} inch.
[0011] In accordance with another aspect of the present invention,
a multi-channel reagent dispenser apparatus for introducing a
plurality of reagents into a plurality of reaction sites comprises
a plurality of reagent sources, and a plurality of reagent
dispensing nozzles each coupled with one of the plurality of
reagent sources. A plurality of dispenser heads each include a
dispenser head body having a dispensing end which is configured to
dispense a plurality of reagents from the plurality of reagent
sources. Each dispenser head body has therein a group of nozzles
being clustered to provide a plurality of nozzle outlets in the
dispenser head body to introduce reagents from the plurality of
reagent sources through the dispensing end of the dispenser head
body into one of the reaction sites. The group of nozzles include
more than one reagent dispensing nozzle from the plurality of
reagent dispensing nozzles.
[0012] In some embodiments, a plurality of reagent dispensing
nozzle valves are each coupled with one of the reagent dispensing
nozzles to control reagent flow from the reagent sources to the
reagent dispensing nozzles. At least one wash nozzle is coupled
with at least one source of wash solvent. The group of nozzles
clustered in each dispenser head body include at least one wash
nozzle. At least one wash nozzle valve is each coupled with one of
the at least one wash nozzle to control wash solvent flow from the
at least one source of wash solvent to the at least one wash
nozzle. The group of nozzles clustered in each dispenser head body
include a vacuum nozzle which is coupled to a vacuum source. A
vacuum nozzle valve is coupled with the vacuum nozzle to control
vacuum flow through the vacuum nozzle.
[0013] In some embodiments, the plurality of dispenser heads
comprise at least one first dispenser head and at least one second
dispenser head. Each first dispenser head has therein a cluster of
first nozzles being coupled with a first set of the plurality of
reagent sources. Each second dispenser head has therein a cluster
of second nozzles being coupled with a second set of the plurality
of reagent sources which are different from the first set of
reagent sources. The first set of reagent sources may comprise
building block elements such as bases A, C, G, T, and an activator
such as tetrazole. The second set of reagent sources may comprise
acid deblock, oxidizers, and capping agents. A first actuator is
configured to move each of the at least one first dispenser head
from one reaction site to another reaction site to introduce
reagents from the first set of reagent sources into the reaction
sites. A second actuator is configured to move each of the at least
one second dispenser head from one reaction site to another
reaction site to introduce reagents from the second set of reagent
sources into the reaction sites. A controller is coupled with the
first and second actuators to automatically control movements of
the at least one first dispenser head and the at least one second
dispenser head to introduce reagents from the first and second sets
of reagent sources separately into the reaction sites.
[0014] In specific embodiments, a plurality of reagent dispensing
nozzle valves are each coupled with one of the reagent dispensing
nozzles. The controller is coupled with the reagent dispensing
nozzle valves to control reagent flow from the reagent sources to
the first nozzles in the at least one first dispenser head and to
the second nozzles in the at least one second dispenser head. The
dispensing end of each dispenser head body has a maximum dimension
of about 9 mm. The plurality of dispenser heads comprise a
plurality of first dispenser heads and a plurality of second
dispenser heads. The first dispenser heads are spaced about 9 mm
apart, and the second dispenser heads are spaced about 9 mm
apart.
[0015] The plurality of reagent sources each are delivered to the
reagent dispensing nozzles by pressurization with an inert gas. The
reaction sites are evacuated under vacuum assist. The reaction
sites are disposed in an array provided in a plurality of vacuum
trays which are formed on a single block. The apparatus is disposed
in an inert environment, such as nitrogen or argon.
[0016] In accordance with another aspect of the present invention,
a method for introducing a plurality of reagents into a plurality
of reaction sites comprises providing a plurality of reagent
sources, a plurality of reagent dispensing nozzles each coupled
with one of the plurality of reagent sources, and a plurality of
dispenser heads. Each dispenser head includes a dispenser head body
having a dispensing end. Each dispenser head body has therein a
group of nozzles being clustered to provide a plurality of nozzle
outlets in the dispenser head body. The group of nozzles include
more than one reagent dispensing nozzle from the plurality of
reagent dispensing nozzles. The method further comprises
controlling flows of reagents from the plurality of reagent sources
through the plurality of reagent dispensing nozzles to dispense a
plurality of reagents via the group of nozzles clustered in each
dispenser head body through the dispensing end of the dispenser
head body into one of the reaction sites.
[0017] In some embodiments, the flows of reagents through each
dispenser head body are controlled by operating a plurality of
reagent dispensing valves each coupled with one of the reagent
dispensing nozzles based on flow rates to dispense preset amounts
of reagents via the group of nozzles clustered in each dispenser
head body at preset times through the dispensing end of the
dispenser head body into one of the reaction sites. The flows of
reagents through each dispenser head body are controlled to provide
one reagent at a time through the dispenser head body.
[0018] The method may further comprise providing at least one
source of wash solvent and at least one wash nozzle coupled with
the at least one source of wash solvent, wherein the group of
nozzles clustered in each dispenser head body include at least one
wash nozzle. The wash solvent is dispensed through the at least one
wash nozzle in each dispenser head body at preset times. The method
may further comprise providing a vacuum nozzle in the group of
nozzles clustered in each dispenser head body, and drawing a vacuum
through the vacuum nozzle in each dispenser head body between
dispensing different reagents through the dispenser head body.
[0019] In some embodiments, the plurality of dispenser heads
comprise at least one first dispenser head and at least one second
dispenser head. Each first dispenser head has therein a cluster of
first nozzles being coupled with a first set of the plurality of
reagent sources. Each second dispenser head has therein a cluster
of second nozzles being coupled with a second set of the plurality
of reagent sources which are different from the first set of
reagent sources. The method further comprises moving each of the at
least one first dispenser head from one reaction site to another
reaction site to introduce reagents from the first set of reagent
sources into the reaction sites. The method may further comprise
moving each of the at least one second dispenser head from one
reaction site to another reaction site to introduce reagents from
the second set of reagent sources into the reaction sites. The at
least one first dispenser head and the at least one second
dispenser head are moved automatically by computer control.
[0020] In specific embodiments, the plurality of dispenser heads
comprise a plurality of first dispenser heads, and flows through
the first nozzles in the first dispenser heads are controlled to
dispense reagents from the first set of reagent sources to separate
reaction sites simultaneously. The plurality of dispenser heads
comprise a plurality of second dispenser heads, and flows through
the second nozzles in the second dispenser heads are controlled to
dispense reagents from the second set of reagent sources to
separate reaction sites simultaneously.
BRIEF DESCRIPTION OF THE DRAWINGS
[0021] FIG. 1 is a simplified schematic diagram of a multi-channel
reagent dispensing system in accordance with an embodiment of the
present invention;
[0022] FIG. 2 is a schematic diagram of a first set of reagent
dispenser heads in the reagent dispensing system of FIG. 1
according to an embodiment of the invention;
[0023] FIG. 3 is a schematic diagram of a second set of reagent
dispenser heads in the reagent dispensing system of FIG. 1
according to an embodiment of the invention;
[0024] FIG. 4 is a sectional view of a reagent dispenser head
according to an embodiment of the present invention;
[0025] FIG. 5 is a schematic diagram of check valve manifolds for
pressurizing the fluid sources in the multi-channel reagent
dispensing system of FIG. 1;
[0026] FIG. 6 is a schematic diagram of a vacuum system in the
multi-channel reagent dispensing system of FIG. 1; and
[0027] FIG. 7 is a schematic diagram of an interface between the
controller and various valves in the multi-channel reagent
dispensing system of FIG. 1.
DESCRIPTION OF THE SPECIFIC EMBODIMENTS
[0028] The present invention is directed to a multi-channel reagent
dispensing system for nuclei acid synthesis or the like. One
embodiment of a reagent dispensing system 10 is shown in the
simplified schematic diagram of FIG. 1. The reagent dispensing
system 10 includes a plurality of reagent and wash solvent sources
12 which are fluidicly coupled with a plurality of dispenser heads
14 to supply solution phase reagents and wash solvents to the
dispenser heads. In the embodiment shown, the dispensing system 10
includes two sets of dispenser heads 14a, 14b, which are fluidicly
coupled with two sets of reagent and wash solvent sources or other
fluid sources 12a, 12b. The first set of dispenser heads 14a are
arranged on a first carrier or support 16a which is driven by a
first actuator 18a, while the second set of dispenser heads 14b are
arranged on a second carrier or support 16b which is driven by a
second actuator 18b. A controller 20 may be provided to
automatically control the actuators 18a, 18b.
[0029] A plurality of vacuum plates or trays 22 are provided, and
each vacuum plate 22 supports a plurality of cells or columns or
reactors 24 which are evacuated under vacuum assist by a vacuum
source coupled to an opening in each vacuum plate 22. In some
embodiments, separate vacuum plates 22 are placed together. In the
embodiment shown in FIG. 1, the vacuum plates 22 are formed on a
single body 26, for instance, by milling them out of a plastic
block. This can save space and improve throughput by providing
vacuum plates 22 that are close together.
[0030] The supports 16a, 16b for the dispenser heads 14a, 14b are
driven by the actuators 18a, 18b to pass over the cells 24, one
cell per dispenser head at a time, and dispense into the cells 24
various fluids including reagents, wash solvents, and the like.
[0031] The two sets of dispenser heads 14a, 14b may be passed over
the cells 24 in different sequences and along different paths. The
controller 20 is desirably a computer controller 20 which is
programmed to move the dispenser head 14a, 14b in desirable
sequences and along desirable paths to achieve the desired
synthesis. The controller 20 may also be coupled with the reagent
and wash solvent sources 16a, 16b to control the dispensing of the
fluids through the dispenser heads 14a, 14b, for instance, by
controlling operation of the valves 32a, 32b (FIGS. 2 and 3)
between the sources 16a, 16b and the dispenser heads 14a, 14b, as
described in more detail below.
[0032] In the embodiment shown, the two sets of dispenser heads
14a, 14b are fluidicly coupled to two different sets of reagent and
wash solvent sources 12a, 12b. In other embodiments, there may be
fewer or more sets of dispenser heads which are fluidicly coupled
to the same or different sets of reagent and wash solvent sources.
The reagent dispensing system 10 is desirably disposed or housed in
an inert environment, such as nitrogen or argon.
[0033] FIGS. 2 and 3 show an example of the two sets of dispenser
heads 14a, 14b fluidicly coupled with two sets of reagents and wash
solvent sources 16a, 16b. In FIG. 2, the first set of fluid sources
16a include bases A (adenine), C (cytosine), G (guanine), T
(thymine), X/Z, TET (tetrazole), and ACN DRY (acetonitrile dry).
The tetrazole may be replaced by another activator. The A, C, G, T,
and X/Z are examples of building block elements, and may be
replaced other building block elements from various molecule types
in other embodiments. Solid phase polymer synthesis methods, for
instance, utilize multiple addition of unit building blocks
including other polynucleic acids (e.g., RNA or DNA/RNA hybrids),
nucleic acid mimics (e.g., PNAs (peptide nucleic acids)), peptides,
and oligosaccharides. The use of the system for combinatorial small
molecule synthesis is also possible.
[0034] The sources 16a each include a plurality of fluid flow lines
30a for separately supplying fluids to each of the dispenser heads
14a. FIG. 2 shows eight channels represented by eight dispenser
heads 14a each containing seven fluid lines 30a from the seven
fluid sources 16a. For clarity, only one dispenser head is shown to
receive fluid lines from all seven fluid sources 16a. Each fluid
line 30a has a corresponding valve 32a that controls fluid flow
from the corresponding fluid source 16a to the dispenser head
14a.
[0035] In FIG. 3, the second set of fluid sources 16b include
DEBLOCK (acid deblock), OX-1 (oxidizer 1), OX-2 (oxidizer 2), ACN
DRUM (acetonitrile drum), CAP A (capping agent A), CAP B (capping
agent B), and OTHER. The sources 16a each include a plurality of
fluid flow lines 30b for separately supplying fluids to each of the
dispenser heads 14b. FIG. 2 shows eight dispenser heads 14b each
containing seven fluid lines 30b from the seven fluid sources 16b.
For clarity, only one dispenser head is shown to receive fluid
lines from all seven fluid sources 16b. Each fluid line 30b has a
corresponding valve 32b that controls fluid flow from the
corresponding fluid source 16b to the dispenser head 14b.
[0036] The use of a separate, dedicated fluid line 30 (30a or 30b)
between each fluid source 16 (16a or 16b) and each dispenser head
14 (14a or 14b) avoids the need for priming the lines, which saves
time and reagents. The nozzles 34 of the fluid lines 30 are
clustered into the dispenser head 14 having an output size
comparable to the size of the cells 24 for dispensing fluids into
the cells 24. FIG. 4 shows a cluster of the fluid nozzles 34 in a
dispenser head 14. In a specific embodiment, each cell 24 has a
diameter of about 9 mm or less, and the dispensing end of each
dispenser head 14 has a maximum dimension of about 9 mm. There are
about eight nozzles 34 clustered in the dispensing head 14, each
nozzle 34 having an outer diameter of less than about {fraction
(1/16)} inch.
[0037] Because each dispenser head 14 has a nozzle 34 for the fluid
line 30 from each fluid source 16 within the set, the set of
dispenser heads 14 need only make one pass over the cells 24 to
supply fluids from the entire set to the cells 24 in a particular
dispensing sequence, thereby improving throughput. The opening and
closing of the valves 32 (32a or 32b) are desirably automatically
performed by computer control. For instance, the valves 32 may be
operatively coupled with the controller 20 which is programmed to
dispense fluids according to a desired synthesis procedure.
[0038] Each set of fluid sources 16 desirably includes a washing
solvent such as acetonitrile. The wash solvent washes the dispenser
heads 14 at various stages during the synthesis procedure to keep
the heads 14 clean. The wash can also minimize or prevent
cross-contamination between the openings of the fluid nozzles 34
clustered within each head 14. As shown in FIG. 4, the nozzle tip
of the wash nozzle 34.sub.W for dispensing acetonitrile or other
wash solvents may be disposed slightly above or proximal of the
tips of the remaining nozzles 34 to ensure that the other nozzles
34 are cleaned by the wash solvent dispensed through the wash
nozzle 34.sub.W. In addition, a vacuum nozzle 34.sub.V is desirably
provided inside the dispenser head 14 to draw out residual fluids
in the dispenser head 14 to minimize or eliminate
cross-contamination. The tip of the vacuum nozzle 34.sub.V is
desirably disposed slight above or proximal of the tips of the
reagent nozzles 34.
[0039] The fluid sources 12 are provided in pressurized containers
or bottles to drive the fluids through the fluid lines 30 to the
dispenser heads 14 by the pressure differential. FIG. 5 shows a gas
source 40 of inert gas such as helium (He) or argon (Ar) used to
pressurize the fluid containers 41. The gas source 40 is coupled to
a plurality of lines having a manual supply valve 42, a pressure
regulator 43, a pressure meter 44, and a flow meter 45. For some
containers 41, multiple-port check valve manifolds are used. FIG. 5
shows, for instance, several 2-port check valve manifolds 46 and an
8-port check valve manifold 47. Of course, other suitable pressure
control and valve arrangements may be used. In other embodiments,
pumps may be used to generate the fluid flow from the fluid sources
to the dispenser heads.
[0040] There are numerous ways to dispense the desired amount of
fluids from the sources 16 into the cells 24. In one embodiment,
the flow from the source 16 to the cell 24 is generated by the
pressure in the pressurized container for the source 16. The flow
rate depends on the number of open valves 32 at that point in time.
The larger the number of open valves 32, the lower the flow rate
and the longer time it takes to keep the valve 32 open to dispense
a given amount of fluid into the cell 24. The controller 20 is
programmed to calculate the flow rate based on the number of open
valves 32 and compute the time required to keep each valve 32 open
to dispense the desired amount of fluid at that flow rate.
[0041] FIG. 6 shows the vacuum system in the multi-channel reagent
dispensing system 10. A vacuum source 50 is coupled with the vacuum
trays 22 (see FIG. 1) via vacuum valves 52 to draw the reagents
through the reaction cells 24 disposed in the vacuum trays 22. A
first set of vacuum valves 52a are provided for the six vacuum
trays 22 on the left side of the body 26, and a second set of
vacuum valves 52b are provided for the six vacuum trays 22 on the
right side of the body 26, in FIG. 1 Any suitable valves may be
used. In the embodiment shown, the vacuum valves 52a and 52b are
pneumatic valves driven pneumatically by a pneumatic source 54 such
as an air source or the like via air or gas manifolds 56a and 56b,
respectively. An example of a suitable valve is a
diaphragm-activated valve available from Parker of Tucson, Ariz.
The vacuum source 50 is further coupled with the vacuum nozzles
34a.sub.V, 34b.sub.V for removing residual reagents in the
dispenser heads 14a, 14b (see FIG. 4), via a first (or left) vacuum
valve 53a for the first set of dispenser heads 14a and a second (or
right) vacuum valve 53b for the second set of dispenser head 14b.
The valves 53a, 53b that are pneumatically driven by the pneumatic
source 54. Vacuum manifolds 58a, 58b are provided for the two sets
of vacuum nozzles 34a.sub.V, 34b.sub.V in each set of dispenser
heads 14a, 14b (see FIGS. 2 and 3).
[0042] The pneumatic source 54 further supplies a gas to a pair of
CAL/VAC (calibration/vacuum) stations 60 associated with the two
sets of dispenser heads 14a, 14b via vacuum valves 55a, 55b. The
CAL/VAC stations 60 are used as waste stations for priming lines
and rinsing nozzle tips, and may also be used to check the flow
rates through the nozzle tips of the nozzles 34 for calibration.
For each set of valves, a pressure sensor 62a or 62b is used to
sense the pressure of the gas provided by the pneumatic source 54,
and a pressure regulator 64a or 64b is used to regulate the
pressure.
[0043] FIG. 7 shows the interface between the controller 20 and
various valves in the multi-channel reagent dispensing system 10.
First I/O boards 70a and a first or left valve driver board 72a are
provided for interface with the valves for the first (or left) set
of dispenser heads 14a. The valves include the dispenser head fluid
line valves 32a for the seven fluid sources (DRY ACN, A, C, G, T,
X/Z, TET) associated with the eight dispenser heads 14a. The seven
fluid sources take up seven ports. An eighth port is used for
vacuum valves, which include vacuum valves 52a (see FIG. 6) for the
six vacuum trays 22 on the left side of the body 26 (see FIG. 1),
the vacuum valve 53a for the first (or left) vacuum nozzles
34a.sub.V, and the vacuum valve 55a for the first (or left) CAL/VAC
station 56a (see FIG. 6). Additional ports may be provided as shown
in FIG. 7. For instance, one port (PORT 8) may provide the
interface for the inert gas (e.g., He) pressure sensor 43 (see FIG.
5), air pressure sensor 62, and other vacuum or volume sensors or
the like.
[0044] Likewise, first I/O boards 70b and a second or right valve
driver board 72b are provided for interface with the valves for the
second (or right) set of dispenser heads 14b. The valves include
the dispenser head fluid line valves 32b for the seven fluid
sources (DRUM ACN, DEBLOCK, OX-1, CAP A, CAP B, OX-2, OTHER)
associated with the eight dispenser heads 14b. The seven fluid
sources take up seven ports. An eighth port is used for vacuum
valves, which include vacuum valves 52b (see FIG. 6) for the six
vacuum trays 22 on the left side of the body 26 (see FIG. 1), the
vacuum valve 53b for the second (or right) vacuum nozzles
34b.sub.V, and the vacuum valve 55b for the second (or right)
CAL/VAC station 56b (see FIG. 6). Additional ports may be provided
as shown in FIG. 7.
[0045] In operation, the controller 20 controls the first actuator
18a to move the first set of dispenser heads 14a across the
reaction cells 24 in each vacuum tray 22 of FIG. 1. The first
actuator 18a aligns the first dispenser heads 14a with the cells 24
for dispensing reagents from the first set of reagent and wash
solvent sources 12a into the cells 24 simultaneously. The type and
amount of reagents dispensed, as well as the delivery of a wash
solvent such as acetonitrile, are controlled by the opening and
closing of the dispenser head fluid line valves 32a by the
controller 20 (see FIG. 7). Typically, one reagent or wash solvent
is dispensed at a time. The controller 20 further controls the
vacuum valve 53a for the vacuum nozzles 34a.sub.V to remove
residual reagents from the dispenser heads 14a and prevent
cross-contamination between different reagents in the dispenser
heads 14a (see FIG. 6). The controller 20 also controls the vacuum
valve 55a for calibration at the CAL/VAC station 50a (see FIG. 6),
and the vacuum valve 52 associated with the particular vacuum tray
22 in which the reaction cells 24 are disposed to receive the
reagents during that time. The first actuator 18a drives the
dispenser heads 14a across the reaction cells 24 in sequence to
dispense the reagents according to desired protocols as programmed
into the controller 20 to achieve the desired synthesis steps in
each of the cells 24.
[0046] The other synthesis steps involve dispensing fluids from the
second set of reagent and wash solvent sources 12b into the
reaction cells 24. The controller 20 controls the second actuator
18b to move the second set of dispenser heads 14b across the
reaction cells 24 in each vacuum tray 22 of FIG. 1. The second
actuator 18b aligns the second dispenser heads 14b with the cells
24 for dispensing reagents from the second set of reagent and wash
solvent sources 12b into the cells 24 simultaneously. The type and
amount of reagents dispensed, as well as the delivery of a wash
solvent such as acetonitrile, are controlled by the opening and
closing of the dispenser head fluid line valves 32b by the
controller 20 (see FIG. 7). Typically, one reagent or wash solvent
is dispensed at a time. The controller 20 further controls the
vacuum valve 53b for the vacuum nozzles 34b.sub.V to remove
residual reagents from the dispenser heads 14b and prevent
cross-contamination between different reagents in the dispenser
heads 14b (see FIG. 6). The controller 20 also controls the vacuum
valve 55b for calibration at the CAL/VAC station 50b (see FIG. 6),
and the vacuum valve 52 associated with the particular vacuum tray
22 in which the reaction cells 24 are disposed to receive the
reagents during that time. The second actuator 18b drives the
dispenser heads 14b across the reaction cells 24 in sequence to
dispense the reagents according to desired protocols as programmed
into the controller 20 to achieve the desired synthesis steps in
each of the cells 24.
[0047] The two actuators 18a, 18b may be controlled to move the
dispenser heads 14a, 14b to different vacuum trays 22 for
dispensing fluids into different cells 24 at different locations in
any desirable sequence. For example, the second dispenser heads 14b
may follow the path of the first dispenser heads 14a, but may also
take on a different path, such as a staggered formation with
respect to the first dispenser heads 14a. Because each set of
dispenser heads 14a, 14b can dispense fluids simultaneously from a
plurality of fluid sources, delivery of reagents and wash solvents
into multiple cells can be achieve without excessive movements and
elaborate control and positioning of the dispenser heads, thereby
improving throughput and reducing the complexity of the delivery
procedure.
[0048] The above-described arrangements of apparatus and methods
are merely illustrative of applications of the principles of this
invention and many other embodiments and modifications may be made
without departing from the spirit and scope of the invention as
defined in the claims. For instance, instead of individual reaction
columns placed within the vacuum tray, the reaction sites may be
formed on a single block and placed in the vacuum tray. The
cross-section of the reaction sites may be round, square, or of
other shapes. In addition, the cells 24 may each have a diameter
greater than 9 mm in some embodiments. In general, the size of the
dispensing end of each dispenser head 14 is smaller than the size
of the cell 24 receiving fluid from the dispenser head 14. For a
standard 96 well microtiter plate, the size is 9 mm. In another
system there may be 20 lines in the nozzle bundle within a single
dispenser head, and the size of the dispensing end of the dispenser
head will be about 12 mm. The reactor cell will have an orifice of
about 14 mm and there will be 48 reactor cells per microtiter
plate. The scope of the invention should, therefore, be determined
not with reference to the above description, but instead should be
determined with reference to the appended claims along with their
full scope of equivalents.
* * * * *